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Exogenous p27KIP1 expression induces anti-tumour effects and inhibits the EGFR/PI3K/Akt signalling pathway in PC3 cells.

ABSTRACT: p27 is a cyclin-dependent kinase inhibitor that regulates the progression of cells from G(1) to S phase of the cell cycle. Loss of p27 has been associated with disease progression and with an unfavourable outcome in prostate cancer. In this study, we investigated whether exogenous p27 expression in the human androgen-independent prostate cancer PC3 cell line had any effect on cell growth, and we studied the molecular mechanisms involved. p27 expression was restored in PC3 cells by plasmid delivery. Cell proliferation and apoptosis were assessed in PC3 cells transfected with p27. We also investigated the effects of p27 on the epidermal growth factor receptor (EGFR)/phosphatidylinositol 3-kinase (PI3K)/Akt signalling pathway in PC3 cells. By restoring p27 expression in PC3 cells, we observed that p27 reduced proliferation and induced arrest in G(0)/G(1) phase. Moreover, p27-transfected PC3 cells underwent apoptosis, as shown by flow cytometric analysis and western blotting analysis of Bcl-2, Bax, Bad, caspase-3 and poly(ADP-ribose)polymerase expression. Furthermore, the p27-induced anti-tumour action correlated with inhibition of the EGFR/PI3K/Akt signalling pathway, as confirmed by western blotting analysis and densitometry of EGFR, PI3K (p85), Akt and p-Akt(S473) expression. Our results suggest that exogenous expression of p27 inhibits the proliferation of PC3 cells through induction of G(1) arrest and apoptosis, and this process correlates with inhibition of the EGFR/PI3K/Akt signalling pathway.

SUBMITTER: Chen J

PROVIDER: S-EPMC3735325 | biostudies-other | 2009 Nov

REPOSITORIES: biostudies-other

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Project description:Background: Oesophageal cancer is the most common malignant tumour with a poor prognosis, and the current treatment methods are limited. Therefore, identifying effective treatment methods has become a research hotspot. Cardamonin (CAR) is a natural chalcone compound and has been reported to play an anticancer role in several cancers. However, its function in oesophageal cancer and the possible underlying mechanism are still unclear. The purpose of this study was to demonstrate the anticancer effect of CAR on oesophageal cancer in vivo and in vitro and to explore the underlying mechanism. Materials and Methods: MTT, crystal violet, and colony formation assays were used to detect oesophageal cancer cell proliferation. The effects of CAR on oesophageal cancer cell migration and invasion were detected by wound healing assay and Transwell assay. Hoechst 33258 staining and flow cytometry were used to detect cell apoptosis. Protein expression levels were detected by Western blot. A tumour xenograft model was established to further test the effect of CAR on the growth of oesophageal cancer in vivo. Results: The results showed that CAR inhibited the proliferation, migration, and invasion of oesophageal cancer cells in a concentration-dependent manner and induced apoptosis. Furthermore, the Western blot assay showed that CAR could suppress metastasis by inhibiting epithelial-mesenchymal transition (EMT) as indicated by downregulated expression of the mesenchymal markers N-cadherin and vimentin, the EMT transcription factor Snail, and matrix metalloproteinases (MMPs) and upregulated expression of the epithelial marker E-cadherin. CAR was associated with upregulation of the pro-apoptotic proteins Bax and Bad and downregulation of the anti-apoptotic protein Bcl-2 and triggered the mitochondrial apoptosis pathway, which in turn promoted caspase-3 activation and subsequent cleavage of PARP; however, the mitochondria-related apoptotic effects induced by CAR were blocked by caspase inhibitor Z-VAD-FMK pretreatment, which prevented programmed cell death triggered by CAR. In addition, CAR reduced the phosphorylation level of downstream effector molecules of phosphatidylinositol 3 kinase (PI3K) in a dose-dependent manner, and treatment with the PI3K agonist 740Y-P could partially reverse the anticancer effect of CAR, demonstrating that CAR played an antitumour role by inhibiting the PI3K/AKT signalling pathway in oesophageal cancer cells. Moreover, the EC9706 xenograft model further confirmed that CAR can significantly inhibit tumour growth in vivo. Conclusion: In summary, CAR exhibited a strong anticancer effect on human oesophageal cancer cells and promoted apoptosis by inhibiting the PI3K/AKT signalling pathway, suggesting that CAR can be used as new strategy for oesophageal cancer treatment.

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Exogenous p27KIP1 expression induces anti-tumour effects and inhibits the EGFR/PI3K/Akt signalling pathway in PC3 cells.

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